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Norovirus detection reagent

a technology of which is applied in the direction of dna/rna fragmentation, biochemistry apparatus and processes, recombinant dna-technology, etc., can solve the problems of not being able to detect other subtypes, norwalk and snow mountain subtype detection reagent,

Inactive Publication Date: 2009-07-16
TOSOH CORP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

The present invention provides a method for detecting all subtypes of norovirus using a gene amplification process. The method involves analyzing the base sequences of norovirus and identifying regions that are common among them. The invention also provides primers that can be used to detect all subtypes of norovirus. The technical effect of the invention is to improve the detectability of norovirus detection and facilitate the development of a reliable and effective reagent for detecting all or most of the subtypes of norovirus.

Problems solved by technology

However, as the primer base sequences are designed on the basis of the base sequence of each subtype, it is not possible to detect other subtypes.
In addition, a norovirus detection reagent using PCR disclosed in Japanese Unexamined Patent Publication No. 2000-300297 can only be used to detect the Norwalk and Snow Mountain subtypes.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

example 1

[0033]In order to show that combination (a) shown in Table 1 is capable of detecting all subtypes of GI and that combinations (b) and (c) are capable of detecting all subtypes of GII, DNA (hereafter, referred to as “artificial standard DNA)” and RNA (hereafter, referred to as “artificial standard RNA”) respectively corresponding to four subtypes of GI and four subtypes of GII were prepared using the methods shown in (1) to (8) below. The artificial standard RNA was measured using the methods shown in (9) to (12).

[0034](1) Artificial standard DNA was designed so as to contain an oligonucleotide binding region used in combinations (a) through (c) based on the base sequence of the Chiba subtype of norovirus (GenBank No. AB042808), and then artificial standard RNA consisting of 90 bases (NV1ART-C, SEQ. ID No. 6) was prepared by in vitro transcription. This standard RNA was used as the sample. After quantifying it by UV absorption at 260 nm, the RNA sample was diluted to 106 copies / 5 μL ...

example 2

[0080]Artificial standard RNA of four GI subtypes were measured according to the methods shown in (1) through (8) below in order to show that one of the combinations of oligonucleotides of the invention of the present application is capable of detecting all subtypes of GI.

[0081](1) The Chiba subtype of artificial standard RNA (NV1ART-C, SEQ. ID No. 6) was used as the sample in the same manner as Example 1. After quantifying it by UV absorption at 260 nm, the RNA was diluted to 106 copies / 5 μL with an RNA diluent (10 mM Tris-HCl (pH 8.0), 1 mM EDTA, 5 mM DTT, 0.25 U / μL RNase inhibitor (Takara Bio)).

[0082](2) The Desert Shield subtype of artificial standard RNA (NV1ART-D, SEQ. ID No. 7) was used as the sample in the same manner as Example 1. After quantifying it by UV absorption at 260 nm, the RNA was diluted to 106 copies / 5 μL with an RNA diluent (10 mM Tris-HCl (pH 8.0), 1 mM EDTA, 5 mM DTT, 0.25 U / μL RNase inhibitor (Takara Bio)).

[0083](3) The Norwalk subtype of artificial standard...

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Abstract

The present invention provides a combination of oligonucleotides preferable for composing a gene testing reagent capable of detecting all subtypes of norovirus rapidly and with high sensitivity. More specifically, the present invention provides a detection method in which only norovirus is specifically amplified and an oligonucleotide that binds to a specific site of norovirus, by using a primer having a sequence that is homologous or complementary to a base sequence specific for norovirus and is located at a position subject to minimal mutation according to subtype.

Description

CONTINUATION DATA[0001]This application is a Continuation of U.S. application Ser. No. 11 / 049,726, filed on Feb. 4, 2005.FIELD OF THE INVENTION[0002]Norovirus is known to be a virus that typically causes viral food poisoning. The present invention relates to a norovirus detection reagent used for detecting norovirus in clinical examinations, public health examinations, food evaluations and food poisoning examinations.PRIOR ART[0003]Norovirus is a member of the human calicivirus family, and has a genome consisting of a single-strand RNA of about 7000 bases. Norovirus is also referred to as Small Round Structured Virus (SRSV).[0004]Roughly 20% of the cases of food poisoning reported in Japan are estimated to be caused by viruses. Norovirus is detected in about 80% of these cases of viral food poisoning. The main infection source is food, and raw oysters are frequently the problem. In addition, norovirus has also been detected in (sporadic) acute gastroenteritis among infants, and the ...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): C12Q1/70C12Q1/68
CPCC12Q1/6883C12Q1/6827C12N15/11C12Q1/6844C12Q1/70C12Q2563/113
Inventor MASUDA, NORIYOSHIYASUKAWA, KIYOSHIHORIE, RYUICHI
Owner TOSOH CORP